Cholesterol pools

In addition to more rapid absorption of lipids in animals fed casein, another mechanism that may be operative is decreased clearance of circulating lipids. Rabbits fed a casein-based semipurified diet excreted significantly less cholesterol but more bile acids in their feces than animals fed a commercial diet (18). The total sterol excretion in feces of the animals fed the casein diet was half that of the rabbits fed the stock diet. Huff and Carroll (19) found that rabbits fed soy protein had a much faster turnover rate of cholesterol and a significantly reduced rapidly exchangeable cholesterol pool compared with rabbits fed casein. Similar studies performed in our laboratory revealed that the mean transit time for cholesterol was 18.4 days in rabbits fed soy protein, 36.8 days in rabbits fed casein, 33.7 days in rabbits fed soy plus lysine, and 36.3 days in rabbits fed casein plus arginine. These data suggest that addition of lysine to soy protein... 

Tissues synthesizing most of the body s cholesterol Cholesterol is synthesized by virtually all tissues in humans, although liver, intestine, adrenal cortex, and reproductive tissues make the largest contribution to the body s cholesterol pool. 

An opposite effect is at the basis of the up-regulation of LDL receptors in response to treatments with bile acid sequestrants, intestinal cholesterol absorption inhibitors, and HMG-CoA reductase inhibitors. The first class of drugs inhibits the intestinal reabsorption of bile acids, thus promoting increased conversion of cholesterol to bile acids in the liver. The increased demand for cholesterol results in activation of the SREBP system and upregulation of LDL receptor synthesis (as well as cholesterol synthesis via upregulation of HMG-CoA reductase). Similarly, inhibition of intestinal cholesterol absorption with ezetimibe results in a reduction in the hepatic cholesterol pool... 

In general, drugs act to reduce the concentration of cholesterol within hepatocytes, causing a compensatory increase in low-density lipoprotein-receptors (LDL-R) on their surface, and increased uptake of cholesterol-rich LDL particles from the bloodstream (see Fig. 25.1). Statins decrease the synthesis of cholesterol and the secretion of VLDL and increase the activity of hepatic LDL-receptors. Bile-acidbinding resins deplete the bile acid and thus the cholesterol pool. Fibrates decrease the secretion of VLDL and increase the activity of lipoprotein lipase, thereby increasing the removal of tri-... 

Cholesterol is formed in the liver (85%) and intestine (12%) - this constitutes 97% of the body s cholesterol synthesis of 3.2 mmol/day (= 1.25 g/day). Serum cholesterol is esterized to an extent of 70-80% with fatty acids (ca. 53% linolic acid, ca 23% oleic acid, ca 12% palmitic acid). The cholesterol pool (distributed in the liver, plasma and erythrocytes) is 5.16 mmol/day (= 2.0 g/day). Homocysteine stimulates the production of cholesterol in the liver cells as well as its subsequent secretion. Cholesterol may be removed from the pool by being channelled into the bile or, as VLDL and HDL particles, into the plasma. The key enzyme in the synthesis of cholesterol is hydroxy-methyl-glutaryl-CoA reductase (HGM-CoA reductase), which has a half-life of only 3 hours. Cholesterol is produced via the intermediate stages of mevalonate, squalene and lanosterol. Cholesterol esters are formed in the plasma by the linking of a lecithin fatty acid to free cholesterol (by means of LCAT) with the simultaneous release of lysolecithin. (s. figs. 3.8, 3.9) (s. tab. 3.8)... 

Rumsey, S. C., Galeano, N., Lipschitz, B., and Deckeibaum, K. J. (1995), Oleate and other long-chain fatty acids stimulate low-density-lipopiotein receptor activity by enhancing acyl coenzyme Aicholesterol acyltransferasc activity and altering intracellular regulatory cholesterol pools in cultured cells. J. Biol. Chem. 270,10006-100016. 

The brain is one of the most cholesterol rich organs of the body and contains 25% of total body cholesterol [92]. While the body manages cholesterol metabolism primarily through the liver, the brain cholesterol compartment is essentially isolated from body cholesterol pools by the blood brain barrier (BBB). During early CNS development, the rate of cholesterol synthesis is quite high due to extensive myelination. As the brain matures, cholesterol synthesis and turnover slow dramatically with an estimated total turnover rate ranging from 4-6 months [93] in adult rat brain. The turnover rate is even slower in humans than in rodents (0.03% per day vs. 0.4% in rodent) [94]. 

Slotte JP, Bierman EL (1988) Depletion of plasma-membrane sphingomyelin rapidly alters the distribution of cholesterol between plasma membranes and intracellular cholesterol pools in cultured fibroblasts. Biochem J 250 653-658... 

In addition to high-affinity receptor-mediated uptake, cultured fibroblasts can take up LDL by receptor-independent bulk fluid pinocytosis (for a review of endocytosis, see ref. 72). LDL uptake by this process is non-saturable and is directly proportional to the concentration of LDL in the medium. Unlike receptor-depen-dent uptake, receptor-independent uptake of LDL is protease-insensitive and does not require divalent cations [28]. In addition, LDL entering cells through non-specific pinocytosis do not increase cellular cholesterol content or regulate cellular cholesterol synthesis or cholesterol esterification [28,73], possibly because the amount of LDL taken up may be insufficient to significantly expand cellular cholesterol pools. 

Exposure of cholesterol-starved cells to LDL is followed by massive buildup of intracellular free cholesterol pools. HMG-CoA reductase is suppressed, shutting down cellular cholesterol synthesis [84,85]. ACAT is stimulated as much as 500-fold by a process that apparently is independent of protein synthesis [86]. Subsequently, the number of LDL receptors declines dramatically with a calculated half-life of 15-20 h [29,31]. Since the rate of decline under conditions which block protein synthesis is comparable to the LDL-mediated rate of decline [29,87], down-regulation may be due to suppression of receptor synthesis. Excess eholesterol is esterified and stored in the c)rtoplasm as cholesteryl esters. A steady-state characterized by large cholesteryl ester and free cholesterol pools and by basal levels of both HMG-CoA reductase and LDL receptors is ultimately attained. This regulatory mechanism allows cells to control their rate of cholesterol uptake, synthesis, and storage in response to the available supply of lipoprotein cholesterol. 

However, foUowing enrichment of the cells with exogenous cholesterol or incubation with 0.1 mM chlorpromazine [156], the whole of the cholesterol pool becomes available to the cholesterol oxidase. This means that the time-course of cholesterol oxidation can be determined and hence the rate of flip-flop. Based on such studies, a half-time of less than 3 sec at 37°C has been found for transposition of cholesterol across the bilayers [157]. These studies have also been extended to probe features of lipid-cholesterol organisation in the human erythrocyte membrane [158]. Clearly, such studies are relevant to our understanding of the mechanisms of cholesterol loss from cells in vivo by the methods outhned earlier. 

Excess cholesterol can also be metabolized to CE. ACAT is the ER enzyme that catalyzes the esterification of cellular sterols with fatty acids. In vivo, ACAT plays an important physiological role in intestinal absorption of dietary cholesterol, in intestinal and hepatic lipoprotein assembly, in transformation of macrophages into CE laden foam cells, and in control of the cellular free cholesterol pool that serves as substrate for bile acid and steroid hormone formation. ACAT is an allosteric enzyme, thought to be regulated by an ER cholesterol pool that is in equilibrium with the pool that regulates cholesterol biosynthesis. ACAT is activated more effectively by oxysterols than by cholesterol itself, likely due to differences in their solubility. As the fatty acyl donor, ACAT prefers endogenously synthesized, monounsaturated fatty acyl-CoA. 

The LDL receptor is a key component in the feedback-regulated maintenance of cholesterol homeostasis [1]. In fact, as an active interface between extracellular and intracellular cholesterol pools, it is itself subject to regulation at the cellular level (Fig. 2). LDL-derived cholesterol (generated by hydrolysis of LDL-bome cholesteryl esters) and its intracellularly generated oxidized derivatives mediate a complex series of feedback control mechanisms that protect the cell from over-accumulation of cholesterol. First, (oxy)sterols suppress the activities of key enzymes that determine the rate of cellular cholesterol biosynthesis. Second, the cholesterol activates the cytoplasmic enzyme acyl-CoA cholesterol acyltransferase, which allows the cells to store excess cholesterol in re-esterified form. Third, the synthesis of new LDL receptors is suppressed, preventing further cellular entry of LDL and thus cholesterol overloading. The coordinated regulation of LDL receptors and cholesterol synthetic enzymes relies on the sterol-modulated proteolysis of a membrane-bound transcription factor, SREBP, as described in Chapter 14. 

Most of these concepts have arisen from detailed studies in cultured fibroblasts from normal subjects and from patients with the disease, FH. Lack of the above-described regulatory features in FH fibroblasts led to the conclusion that the abnormal phenotype is caused by lack of LDL receptor function, and thus, disruption of the LDL receptor pathway. In particular, the balance between extracellular and intracellular cholesterol pools is disturbed. Clinically, the most important effect of LDL receptor deficiency is hypercholesterolemia with ensuing accelerated development of atherosclerosis and its complications (Chapter 21). In the following sections, a detailed description of the... 

Ann Jeina was treated with cholestyramine, a resin that binds some of the bile salts in the intestine, causing these resin-bound salts to be carried into the feces rather than recycled to the liver. The liver must now synthesize more bile salts, which lowers the intrahepatic free cholesterol pool. As a result, hepatic LDL receptor synthesis is induced, and more circulating LDL is taken up by the liver. 

HMG-CoA reductase inhibitors, such as pravastatin, also stimulate the synthesis of additional LDL receptors but do so by inhibiting HMG-CoA reductase, the rate-limiting enzyme for cholesterol synthesis. The subsequent decline in the intracellular free cholesterol pool also stimulates the synthesis of additional LDL receptors. These additional receptors reduce circulating LDL-cholesterol levels by increasing receptor-mediated endocytosis of LDL particles. 

Ann Jeina was treated with a statin (pravastatin) and cholestyramine, a bile acid sequestrant. With the introduction of the cholesterol absorption blocker ezetimibe, the use of cholestyramine with its high level of gastrointestinal side effects may decline. Ezetimibe reduces the percentage of absorption of free cholesterol present in the lumen of the gut and hence the amount of cholesterol available to the enteroc5de to package into chylomicrons. This, in turn, reduces the amount of cholesterol returning to the liver in chylomicron remnants. The net result is a reduction in the cholesterol pool in hepatocytes. The latter induces the synthesis of an increased number of LDL receptors by the hver cells. As a consequence, the capacity of the liver to increase hepatic uptake of LDL from the circulation leads to a decrease in serum LDL levels. 

Not all brain cholesterol is, however, metabolically stable, for some turnover has been demonstrated in the gray matter of young rabbits (Davison et al, 1959a). Other experiments by Kabara and Okita (1961) also indicate that there are several sterol compartments in brain, each with a difiEerent turnover rate. Very rapid labeling of a small cholesterol pool followed injection of or H -labeled precursors into young adult mice at a rate equivalent to almost complete turnover in 80 minutes. Pritchard (1963) also found catabolism of 50% of C -Iabeled cerebral cholesterol 30 days after injection of 1-C -acetate into 1-day-old rats, although during the remainder of the experimental period (40 days) the radioactive cholesterol was retained in the brain with little loss. 

There is no direct evidence so far that the actual concentration of serum cholesterol would determine bile acid production and elimination in man. For instance, increase of serum cholesterol by dietary cholesterol is not associated with compensatory increase in bile acid production (63,71,86,87). This does not exclude the possibility that an increase of some lipoprotein subfraction would stimulate bile acid synthesis. Thus determinations of bile acid synthesis by the isotope dilution method have shown markedly high values in triglyceridemic subjects (69), though according to sterol balance data this association is mostly determined by the degree of obesity of these patients (11,63). It is also interesting to note that though the serum cholesterol level and bile acid production are not normally correlated with each other, bile acid synthesis and the serum cholesterol pool are closely correlated in normocholesterolemic nonobese and obese subjects and in hypercholesterolemic individuals (88). 

The negative correlation between fecal bile acids and serum cholesterol was of low degree for the total series (r = —0.24) and for nonobese patients (r = —0.27), the correlation being positive with the serum cholesterol pool (r = 0.28). Fecal bile acids also correlated positively with body weight (r = 0.52), relative body weight (r = 0.31), and body surface (r = 0.53). 

In rodents taken as animal models, the cholesterol pool of adipose tissue is present in unesterified form and localized to the bulk triglyceride storage droplets. The cholesterol content increases with age, diet, and state of obesity. 

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